We intend to probe enzymatic reaction mechanism from three separate but complementary routes. First we shall probe enzymatic catalyses with suicide substrates which are chemically unreactive in solution but become activated only at the active site of specific target enzymes which, recognizing them as substrates, transform them to intermediates where latent reactive functionalities can become unmasked and lead to covalent active site blocking. In addition to providing information on catalytic pathways and the identification of functional active rite components, these analogs offer sufficient selectivity to offer eventual therapeutic value in vivo. By using acetylenic substrates and oxidizable alkenoic dehydrogenases, pyridoxal and coenzyme B12 dependent lysine metabolizing enzymes, transaminases, and various racemases and isomerases. We intend to examine through stereochemical and other mechanistic analysis two unusual reactions carried out by the flavoenzyme D-amino acid oxidase and the PLP-folate dependent serinehydroxymethylase. In the first instance we may gain insight into how flavoenzymes modulate reactivity of reduced coenzyme with molecular oxygen by study of an alpha, beta-elimination on a beta-halosubstrate. In the second case, experiments may indicate an unusual lack of stereochemical control by the serine enzyme and offer insight into the role of tetrahydrofolate as aldehyde carrier. BIBLIOGRAPHIC REFERENCES: Ghisla, S., Ogata, H., Massey, V., Schonbrunn, A., Abeles, R. and Walsh, C. Kinetic Studies on the Inactivation of L-Lactate Oxidase by the Acetylenic Suicide Substrate 2-Hydroxy-3-Butynoate. Biochemistry 15: 1791 (1976). Schonbrunn, A., Abeles, R., Walsh, C., Ghisla, S., Ogata, H. and Massey, V. The Structure of the Covalent Adduct Formed Between Lactate Oxidase and the Suicide Substrate 2-Hydroxy-3-Butynoate. Biochemistry 15: 1798 (1976).